We propose to analyze the role of oncogenes in the pathogenesis of human melanoma. Our recent work has shown that 4 out of 40 (10%) cultured human melanoma cell lines have activated transforming genes detectable in the NIH/3T3 assay. Of the 40 melanoma lines assayed, five were from separate metastatic deposits from a single patient. Only 1 of the 5 cell lines from this patient had detectable oncogene activity. The transforming genes derived from the four positive melanoma cell lines were found to belong to the ras gene family, with three isolates related to the N-ras gene and one isolate related to the c-Ha-ras-1 gene. To extend these initial studies, we propose to: (1) determine the frequency of ras gene activation in non-cultured primary and metastatic deposits from different individuals, and in separate multiple primary and metastatic deposits from the same individual. Further, we will investigate p21 levels, p21 species and p21 message in a panel of cultured melanoma cells; (2) determine the presence of transforming non-ras oncogenes in non-cultured and cultured primary and metastatic melanomas; (3) determine the pattern of oncogene expression in cultured and non-cultured primary and metastatic melanomas; and (4) develop transformation assays using human melanocytes and nevus cells. Initial studies will determine optimal conditions for DNA uptake and stable expression and if these cells are competent for transformation by infection with SV40 virus. We will assay the transforming activity of the following cloned genes alone or in combinations: human c-myc, human Ha-ras and adenovirus early region 1A. Studies will also be undertaken to test the transforming activity of human N-ras and c-myc genes cloned into a retroviral experssion vector, pZipNeoSV(X). (X)